Prostate carcinoma is the most common cancer diagnosed in men in the United States and has the second highest cancer death rate yielding only to lung adenocarcinoma. Parker et al., CA Cancer J. Clin. 46:5-27 (1996). Although it is possible to effectively treat organ-confined prostate cancer, there are very limited treatment options for metastatic disease. Thus, it is of great importance to find novel ways to diagnose early stage disease and to closely monitor both progression and treatment of the disease, as well as to develop new therapeutic approaches. To achieve this, it is important to understand the molecular mechanisms of prostate cancer development and to identify new biochemical markers for disease diagnosis and progression.
To date there are very few prostate-specific markers available. The best-known and well-characterized markers of proven prostate cancer diagnostic value are the proteins prostatic acid phosphatase (PAP), prostate specific antigen (PSA), and prostate-specific membrane antigen (PSMA). Each of these proteins has also become the target for novel immunotherapy approaches to the treatment of disease. Horoszewicz et al., Anticancer Res. 7:927-935 (1987); Barren et al., Prostate 30:65-68 (1997); Murphy et al., Prostate 33:281-285 (1997); Murphy et al., Prostate 26:164-168 (1995); Rochon et al., Prostate 25:219-223 (1995); Correale et al., J. Immunol. 161:3186-3194 (1998); and Murphy et al., Prostate 38:73-78 (1999).
It has been reported that a cation channel protein, variously designated Trp-p8 (transient receptor potential-p8), TRPM8, and CMR1 (cold and menthol receptor 1), is preferentially expressed in prostate. Cloning of the full-length human trp-p8 cDNA revealed a transcript corresponding to an 1104 amino acid polypeptide sharing homology with the trp family of calcium channels. Clapham et al., Nature Reviews 2:387-396 (2001) and Clapham et al., IUPHAR Compendium, TRP Channels (2002). Trp-p8 shows particularly high homology with the human TRPC7 gene—a putative Ca2+ channel protein of the trp family that is highly expressed in brain tissue. Nagamine et al., Genomics 54:124-131 (1998). Trp-p8 also shows significant homology to human melastatin, another Trp family-related protein expressed in melanocytes and believed to be a tumor suppressor gene. Duncan et al., Cancer Res. 58:1515-1520 (1998); and Hunter et al., Genomics 54:116-123 (1998). Perhaps of greatest interest is the observation that the trp-p8 gene appears to be expressed in a large spectrum of nonprostatic in addition to prostatic neoplastic lesions. Tsavaler et al., Cancer Res. 61(9):3760-9 (2001).
The Trp superfamily comprises more than 20 related cation channel proteins that have been implicated in processes including sensory physiology to vasorelaxation and male fertility. Defects in Trp channels have been associated with changes in growth control and tumor suppression. While all Trp proteins are calcium channels, they vary significantly in their selectivity and mode of activation. Members of the Trp superfamily share significant sequence homology and predicted structural similarities, such as size of predicted transmembrane segments.
Trp-p8 is over-expressed in a range of cancers including prostate, breast, lung and colon, while within normal tissues, it is predominantly expressed in human prostate [Tsavaler et al., supra] and dorsal root ganglia (DRG, Dendreon, unpublished observation). Fuessel et al. reported that Trp-p8 is a highly prostate-specific and prostate carcinoma-associated gene thus qualifying it as a potential target for specific therapies. International J. of Oncology 23:221-228 (2003). Among other species, Trp-p8 orthologues are reportedly expressed in a subset of DRG and trigerminal ganglia (TG) neurons in rat [McKemy et al., Nature 416(6876):52-8 (2002)] and mouse [Peier et al., Cell 108(5):705-15 (2002)] as well. Thus, Trp-p8 is a pantumor-expressed marker with significant potential use in disease diagnosis and monitoring of disease progression during treatment as well as a viable target for cancer therapy.
In two articles published concurrently, it was reported for the first time that Trp-p8 orthologues, in response to cold and certain cooling compounds, initiate an influx of cations, such as calcium, from the extracellular space. McKemy et al., supra; and Peier et al., supra. Two of the best known modulators of Trp-p8 activity are the Trp-p8 agonists menthol and Icilin. Menthol is effective in inducing calcium influx at ˜10-100 μM while Icilin is more potent with an effective concentration range of 0.1-1 μM.
The higher temperature threshold reported for Trp-p8 activation by the most widely studied agonist, menthol (2-isopropyl-5-methyl-cyclohexanol), is about 30-32° C. in a variety of cells (cold-sensitive neurons, Trp-p8 heterologously expressed in Xenopus oocytes, HEK293 and CHO cells). McKemy et al., Nature, supra; Peier et al., Cell, supra; Nealen et al., J Neurophysiol. 90(1):515-520 (2003); and Reid et al., J Physiol. 545(Pt 2):595-614 (2002).
Although certain agonist compounds have been shown to activate Trp-p8 expressing cells up to 32° C., there has been no report disclosing growth manipulation in cells at physiological temperature (i.e. 37° C.), the temperature at which a compound must be active in order to be an efficacious in vivo therapeutic.
Association of Trp-p8 with prostate, lung, breast, and colon cancers and the important role various ion channels play in vital cell functions suggest that Trp-p8 channel may have a significant function in cancer cell signaling and/or proliferation. Modulation of Trp-p8 activity, either by activating via an agonist or inhibiting via an antagonist, at a physiological temperature can be valuable as a therapeutic to manipulate the Trp-p8 expressing cells in a specific manner.
Accordingly, there remains a need in the art for small-molecule modulators of Trp-p8 activity, compositions comprising one or more small-molecule Trp-p8 modulators, and methods for the identification and use of small-molecules for modulating the activity of Trp-p8 in a cell and for the treatment of disease associated with the aberrant expression of Trp-p8.